PROJECT SUMMARY/ABSTRACT The focus of this proposal is to generate ?humanized? models of AD in Drosophila with robust AD-relevant phenotypes and utilize them to identify proximal causes of AD, reliable biomarkers of disease progression, and discover novel avenues to treatments. Unlike transgenic over-expression models, our ?humanized? genetic model will convert the endogenous fly dAPPL, psn, and dBACE genes to their human counterparts. This will preserve all of the complexities of human APP metabolism in the fly by recapitulating key aspects of human AD in vivo in this experimentally malleable model organism, providing a unique and valuable platform for the discovery of new and novel genetic and pharmacological interventions for the treatment of AD. A systems biology approach will be employed to integrate gene expression and metabolomics with relevant AD phenotypes using these genetic models to facilitate our understanding of the proximate causes of AD and for the development of biomarkers of disease progression. Finally, these new ?humanized? AD models will be used in powerful forward genetics screens in Drosophila for suppressors of AD disease to directly identify novel proteins and pathways that reveal new avenues to effective therapies for the treatment of AD. The aims of this proposal are to: (i) develop new ?humanized? models of AD in the fly by humanizing the fly cognate genes to be dh(drosophila-to-human)APP, dhPSN and dhBACE, and introducing a series of known human familial early onset AD mutations in these humanized genes to perturb the endogenous production of human A? in flies to create fly models with robust AD-relevant phenotypes; (ii) assess genetic interactions between humanized loci for dhAPP, dhPSN and dhBACE to determine whether certain combinations of human AD mutant alleles achieve maximal A? production and accelerate phenotypes; (iii) utilize systems biology approaches to integrate transcriptomics and metabolomics with disease phenotypes to determine the molecular and genetic pathways associated with disease progression and to develop reliable biomarkers of disease progression; and (iv) make use of the enormous power of fly genetics to perform an unbiased forward genetic suppressor screen on the most robust AD genetic model to identify new targets for treatment of AD. To better understand the proximal molecular events leading to the progression to neurodegeneration and dementia in AD and to identify new interventions for its treatment , we will develop: (i) new humanized fly models for AD in which flies will express normal and mutated human proteins from AD-relevant cognate fly genes in order to replicate the important pathological processes of human AD in Aim 1; (ii) utilize RNA-seq and metabolomics to measure the transcriptome and metabolome of the humanized AD models and employ systems biological approaches to integrate these data with AD phenotypes in Aim 1 and 2; (iii) perform an unbiased forward genetic screen for suppressors of the robust human AD genetic models in the fly in order to identify proteins and pathways involved in AD in Aim 3.